1. Field of the Invention
Three related compounds, 11-hydroxy-7-methoxy-camptothecin (11,7-HMCPT), 11-hydroxy camptothecin (11-HCPT) and 11-hydroxy 7-ethyl camptothecin (11,7-HECPT), which have previously demonstrated antitumor activity, have not yet been further developed due to poor water solubility. Additionally, these compounds have not been further developed because of structural modifications due to synthetic inefficiency in the preparation of 11,7 substituted camptothecins using shorter synthetic schemes than total synthesis. This invention overcomes these limitations and claims novel compositions of matter, pharmaceutically acceptable formulations of 11,7-HECPT and 11,7-HMCPT and antitumor compositions comprising 11,7-HECPT and 11,7-HMCPT. Additionally, this invention claims novel dosages, schedules of administration, and routes of administration for both 11,7-HECPT and 11,7-HMCPT formulations to humans with various forms of cancer.
2. Description of the Related Art
A. Introduction
Camptothecin (CPT) is highly water insoluble. The creation of water soluble derivatives of camptothecin has been the subject of pursuit by many investigators in order to simplify or eliminate several significant technical problems associated with administration of this class of highly water insoluble drugs.
One water soluble derivative of camptothecin is CPT-11. CPT-11 has undergone Phase I and Phase II clinical trials in human patients with cancer. CPT-11 is a product which requires metabolic conversion to an active species. Metabolic conversion of CPT-11 (a water soluble derivative of camptothecin) to its active metabolite, 10-hydroxy-7-ethyl camptothecin (SN38), varies from patient to patient. This required conversion of CPT-11 to SN38 limits the utility of CPT-11 because of the difficulty in reliably and safely achieving the highest tolerated plasma concentrations of SN38 in the patient.
SN38 is poorly soluble in water. The conversion of CPT-11 to SN38 involves a putative carboxyl esterase enzyme, which is believed to be mainly responsible for the metabolic production of SN38 from CPT-11. Human lung cancer cell lines have been observed to convert less CPT-11 to SN38 than normal cells. The cancer cells' decreased metabolic conversion represents a form of resistance to CPT-11 and limits the utility of CPT-11 in terms of reliably and safely achieving adequate plasma concentrations of SN38 to inhibit the growth of cancer cells in humans. 11-hydroxy-7-alkoxy camptothecins, 11-hydroxy-7-alkyl camptothecin represent another new class of antitumor compounds. These compounds do not require metabolic activation to produce active species, and like camptothecin are highly water insoluble.
The present invention, using both 11,7-HECPT and 11,7-HMCPT, takes advantage of the high lipid solubility and the lack of metabolic activation along with formulation art which permits direct administration of these camptothecin derivatives to human patients with cancer.
Until now, 11,7-HECPT and 11,7-HMCPT derivatives of camptothecin have not been described because of prior art which teaches that water insolubility generally renders alkyl substituted camptothecins unsuitable for direct clinical use because of limitations in pharmaceutical formulations.
This invention teaches the formulation of two specific camptothecin (CPT) derivatives, namely 11-hydroxy-7-methoxy camptothecin (11,7-HMCPT) and 11-hydroxy-7-ethyl camptothecin (11,7-HECPT) in a pharmaceutically acceptable manner using an organic solvent or a mixture of organic cosolvents which permit direct administration of these species of CPT to cancer patients.
This invention also provides certain indications, schedules, dosages and routes of administration for 11,7-HECPT and 11,7-HMCPT for the purpose of treating cancer in humans. The selection of suitable organic solvents for pharmaceutical dosage formulations of the claimed invention is limited to organic solvents with a high degree of physiological safety.
Additionally, this invention teaches administration of 11,7-HECPT or 11,7-HMCPT in a pharmaceutically acceptable multi-solvent formulation. The claimed formulations overcome interpatient variability and biochemical drug resistance which are associated with the use of CPT-11.
The claimed invention also teaches the use of 11,7-HECPT and 11,7-HMCPT in treating human patients with cancer where the cancer cells may have become resistant to metabolic conversion of CPT-11 to SN38 because of altered enzymatic activity.
B. DNA Topoisomerases
Several clinically important anticancer drugs kill tumor cells by affecting DNA topoisomerases. Topoisomerases are essential nuclear enzymes that function in DNA replication and tertiary structural modifications, such as overwinding, underwinding, and catenation, which normally arise during replication, transcription, and perhaps other DNA processes. The two major topoisomerases that are ubiquitous to all eukaryotic cells are topoisomerase I (topo I), which cleaves single stranded DNA, and topoisomerase II (topo II), which cleaves double stranded DNA. Topoisomerase I is involved in DNA replication; it relieves the torsional strain introduced ahead of the moving replication fork.
Topoisomerase I, purified from human colon carcinoma cells or calf thymus, is inhibited by camptothecin, a water soluble analog (CPT-11), and its proposed active metabolite, 10-hydroxy-7-ethyl camptothecin (also known as SN38). Another water soluble Topo I inhibitor in clinical trials is topotecan. These camptothecin derivatives inhibit the enzyme by an identical mechanism; they stabilize the covalent complex of enzyme and strand-cleaved DNA, which is an intermediate in the catalytic mechanism. The compounds have no binding affinity for either isolated DNA or topoisomerase I but bind with measurable affinity to the enzyme-DNA complex. The stabilization of the topoisomerase I "cleavable complex" by camptothecins with intact lactone E-ring is readily reversible. Although camptothecins generally have no effect on topoisomerase II, camptothecin, CPT-11 and SN38 stabilize the "cleavable complex" in a manner analogous to the way in which epipodophyllotoxin glycosides and various anthracyclines inhibit topoisomerase II.
Inhibition of topoisomerase I by camptothecin induces protein-associated-DNA single strand breaks. Virtually all of the DNA strand breaks observed in cells treated with certain camptothecin derivatives are protein linked, whereas an increase in unexplained protein-free breaks can be detected in L1210 cells treated with camptothecin. The compounds appear to produce identical DNA cleavage patterns in end-labeled linear DNA. Under no circumstance has it been demonstrated that camptothecin, CPT-11, SN38, or topotecan cleaves DNA in the absence of the topoisomerase I enzyme.
C. Activity of Camptothecin, HECPT, HMCPT, Topotecan and CPT-11 is Cell Cycle Specific
The antitumor activity of camptothecins is cell cycle specific. The greatest quantitative biochemical effect observed in cells exposed to camptothecin is DNA single strand breaks that occur during the S-phase. Because the S-phase is a relatively short phase of the cell cycle, longer exposure to the drugs results in increased cell killing. Brief exposure of tumor cells to the drugs produces little or no cell killing, and quiescent cells are refractory. These results are likely due to two factors:
(1) The drugs inhibit topoisomerase I reversibly. Although they may produce potentially lethal modifications of the DNA structure during DNA replication, the breaks may be repaired after washout of the drug; and PA1 (2) Cells treated with topo I inhibitors such as camptothecin tend to stay in G.sup.0 of the cell cycle until the drug is removed and the cleaved DNA is repaired. Inhibitors of these enzymes can affect many aspects of cell metabolism including replication, transcription, recombination, and chromosomal segregation. PA1 Sub crop: Repeated recovery of a desired product via pooling of an impure mixture obtained from the first batch of fractional separation. PA1 Mother liquor: The main solvent portion and subsequent washings that contain the desired product. The mother liquor becomes contaminated with impurities during recrystallization step. PA1 Soxhlet Extractor: A glass apparatus used for the effective separation of a material that has preferential solubility in a particular solvent as compared to the rest of the components in a mixture. PA1 Volatile carboxylic acid: This acid is a low boiling organic acid. Examples of a volatile carboxylic acid are formic acid, acetic acid, trifluoroacetic acid, and propionic acid. PA1 Quartenization: This process describes how weak organic bases, such as tertiary nitrogen atoms, can be converted to corresponding salts ionic in nature by treating a weak organic base with either a carboxylic acid or a mineral acid. PA1 Examples of chromatography: Examples of chromatography include: Silica gel chromatography, cellulose chromatography, ion-exchange chromatography, reverse phase chromatography, gel filtration chromatography, and florisil chromatography. For the purpose of this invention, the inventors prefer to use silica gel chromatography. PA1 Lower case alcohols: Lower case alcohols are alcohols that boil below 125.degree. C. Examples of lower cases alcohols are methanol, ethanol, propanol, isopropanol, butanol, 2-butanol, isobutanol, and tertiary butanol. PA1 (1 )direct administration of 11,7-HECPT or 11,7-HMCPT allows the clinician to tailor administration of active cytotoxic species to suit the patient's tolerance; PA1 (2) direct administration of 11,7-HECPT or 11,7-HMCPT overcomes interpatient variability which may be due to polymorphism of key enzyme(s) in the metabolism of water soluble product forms of camptothecin such as CPT-11; and PA1 (3) clinicians can more consistently optimize the drug dosage and schedule to achieve the maximum tolerated dose of 11,7-HECPT or 11,7-HMCPT which is likely to lead to the most beneficial clinical anti-cancer effect. PA1 (1) methods of administering 11,7-HECPT or 11,7-HMCPT to patients with cancer; PA1 (2) solutions of 11,7-HECPT or 11,7-HMCPT; PA1 (3) antitumor compositions comprising 11,7-HECPT or 11,7-HMCPT; PA1 (4) stable formulations of 11,7-HECPT or 11,7-HMCPT suitable for parenteral administration; PA1 (5) pharmacologic schedules for achieving the maximum tolerated dose with acceptable clinical toxicity observed in standard clinical practice of cancer treatment (&lt;10% Grade 3 Toxicity by WHO Classification); PA1 (6) a novel oral formulation of 11,7-HECPT or 11,7-HMCPT; and PA1 (7) use of 11,7-HECPT or 11,7-HMCPT for the treatment of localized complications of cancer by direct administration via instillation into various body cavities.
D. Lactone Form Stabilizes Antitumor Activity of Camptothecin and its Derivatives with the Native E Ring.
Researchers have demonstrated by HPLC, NMR and other techniques that camptothecin, topotecan, CPT-11, and other camptothecin derivatives undergo an alkaline, pH-dependent hydrolysis of the E-ring lactone. The slow reaction kinetics allow one to assess whether both the lactone and non-lactone forms of the drug stabilize the topoisomerase I-cleaved DNA complex. Studies indicate that only the closed lactone form of camptothecin helps stabilize the cleavable complex. This observation provides some rationale for the high degree of activity observed in solid tumor models with exposure to camptothecin. Tumor cells, particularly hypoxic cells prevalent in solid neoplasms, have lower intracellular pH levels than normal cells. At pH levels below 7.0, the lactone form of camptothecin predominates. Thus, one would predict that the drug will be more effective at inhibiting topoisomerase I in acidic cells than in cells having higher intracellular pH levels.
E. Camptothecin, CPT-11 and Topotecan
Wall and Wani isolated camptothecin from the plant, Camptotheca acuminata, in 1966. In the early 1970's, camptothecin reached Phase I trials and was found to have some antitumor activity, but it caused unpredictable myelosuppression and hemorrhagic cystitis. Phase II studies with sodium camptothecin were limited because they induced unpredictable and severe myelosuppression, gastrointestinal toxicity, hemorrhagic cystitis, and alopecia. Clinical trials with sodium camptothecin were eventually discontinued because of these unpredictable toxicities and the lack of significant anti-tumor activity.
Two camptothecin derivatives, CPT-11 and topotecan, have less sporadic toxicities but retain the significant antitumor activity of the parent compound. CPT-11 and topotecan are undergoing Phase I and Phase II development in the United States. 10,11-methylene dioxycamptothecin (MDCPT) is reportedly very active in preclinical studies, but it is also reported to be relatively insoluble in water which reportedly limits its use in the clinic.
Tables 1 and 2 present data summarizing Phase I and Phase II clinical trials of CPT-11. Neutropenia and diarrhea are the major reported, dose-limiting toxicities of CPT-11.
TABLE 1 __________________________________________________________________________ PHASE I STUDIES OF CPT-11 # Investig ator Schedule Pts Dose Toxicity Tumor Type __________________________________________________________________________ Clavel et al 90 min. 37 115 mg/m2/d Neutropenia* Breast (1 PR) QDx 3 Q21 (33-115) diarrhea, Mesothelioma days nausea and (1 PR) vomiting, alopecia Culine et al 90 min. 59 150 mg/m2/wk Neutropenia* esophagus Q21 days (50-150) diarrhea* (1 PR) cervix vomiting, (1 PR) renal alopecia (1 PR) ovarian fatigue (1 PR) stomatitis Neutropenia* Negoro et al 30 min 17 100 mg/m2 Diarrhea*, N/V, NS CLC (2 PRs) infusion (50-150) alopecia, weekly liver dysfunction Ohe et al 120 hr Cl 36 40 mg/m2/d Diarrhea* None Q3 wks (5-40) nausea and vomiting, thromobo- cytopenia, anemia, liver dysfunction Diarrhea* Rothenberg 90 mg QWx 4 32 180 mg/m2/wk Neutropenia, Colon Ca (2 PRs) et al Q42 days (50-180) nausea, vomiting, alopecia Rowinsky 90 min 32 240 mg/m2 Neutropenia* Colon Ca (1 PR) et al infusion (100-345) vomiting, Cervix Ca Q21 day diarrhea abd. (1 PR) pain, flushing __________________________________________________________________________ *Dose Limiting Toxicity .sub.-- Dated updated, unpublished
TABLE 2 __________________________________________________________________________ CPT-11 PHASE II TRIALS INVESTIGATOR TUMOR TYPE SCHEDULE # Pts. RESPONSE __________________________________________________________________________ Fukuoka et al Untreated 100 mg/m.sup.2 weekday 73 (23/72) Neutropenia diarrhea, Non Small PRs nausea, vomiting, Cell Lung 31.9% anorexia, alopecia Cancer Masudu et al Refractory or 100 mg/m.sup.2 weekly 16 (7/15) Neutropenia, diarrhea, Relapsed PRs pneumonitis Small Cell 47% (12.5%) Lung Cancer Negoro et al Small Cell 100 mg/m.sup.2 /week 41 2 CRs Neutropenia (38.6%) Lung Cancer and 7 N/V (61.5%) PRs diarrhea (53.8%) 33.3% alopecia (40.0%) Ohono et al Leukemia/ 200 mg Q3 No resp. 62 ** Neutropenia (91%) Lymphoma 40 mg/m.sup.2 Q0x5 Thromocytopenia 34% PR Gastrointestinal (76%) 20 mg/m.sup.2 bid x7 25% RR Shimada et al Colon cancer 100 mg/m.sup.2 /week or 17 6/17(PR) Neutropenia (53%) 150 mg/m.sup.2 /Q 2 wks 46% N/V (35%) diarrhea (24%) Takeuchi et al Cervical 100 mg/m.sup.2 weekly 69 SCR Neutropenia (89%) cancer 150 mg/m.sup.2 weeks 8 PR N/V (51%) RR of Diarrhea (39.1%) 23.6% Alopecia (38.1%) __________________________________________________________________________
F. Water Insoluble SN38 is the Active Metabolite of CPT-11
Preclinical data obtained from animals and more recent data obtained from humans by Barilero et al. suggest that SN38 is the active metabolite of CPT-11 in vivo. Several different researchers administered CPT-11 intravenously during Phase I trials and recorded the peak plasma concentrations (CpMax) at the end of the infusions. An analysis of the published mean peak plasma concentrations indicates that approximately 1.5% to 9% of the administered CPT-11 (on a per/mg basis) is converted into SN38. The pharmacokinetic data from 30-minute intravenous infusions show a lower percentage of conversion (approximately 1.5%) of CPT-11 to SN38 than that observed following more prolonged infusions (approximately 9% at 40mg/m.sup.2 /d.times.5). The reported half life of SN38 observed in humans following the administration of CPT-11 ranges from 8.8 to 39.0 hours.
The biochemical and pharmacologic relationship between CPT-11 and SN38 and the role of theses compounds in killing cancer cells in vivo is not completely understood. Investigators looking at tumor cell lines in vitro have reported that SN38 has 3600-fold greater inhibitory activity than CPT-11 against topoisomerase I enzyme of P388 cells and that SN38 is approximately 1000-fold more potent in generating single-strand DNA breaks in MOLT3 cells. However, Kaneda et al. report that SN38 has little anti-tumor activity compared to CPT-11 in vivo. They base their findings on studies conducted using an intermittent bolus schedule (days 1, 5, and 9) and an intraperitoneal route of administration with an intraperitoneal P388 tumor model in mice. The inventors believe that the activity of SN38 would have been observed to be greater had these investigators increased the duration of drug exposure instead of using an intermittent bolus schedule.
Ohe et al. suggest that SN38 is a more toxic moiety of CPT-11 and could be responsible for much of the toxicity attributed to CPT-11. However, these same investigators noted a lack of correlation between SN38 pharmacokinetics and dose or CPT-11 pharmacokinetics and toxicity in human subjects. Furthermore, Ohe et al. noted a large range of interpatient variability in the AUC of CPT-11 and its metabolism to SN38, which may result in unpredictable variability in the pharmacokinetic behavior, clinical anti-tumor effects, and toxicity in the individual patient. The data Ohe et al. obtained (using a 5-day, continuous intravenous infusion of CPT-11) also suggests that the conversion of CPT-11 to SN38 is a saturable process. If this is so, the clinical approach to maximizing dose intensity of the SN38 active metabolite would impose additional limitations on the effective use of CPT-11 in patients with cancer.
In preclinical studies of xenografts of human tumors in nude mice, Kawato et al. report that the sensitivity of human tumors to CPT-11 is independent of their ability to produce SN38 and that the effectiveness of CPT-11 is not related to the ability of the tumor to produce SN38. The Kawato et al. report suggests further that SN38 production is likely to be mediated in the plasma or interstitial compartment. Kaneda et al. observed that the plasma concentration of SN38 in mice was maintained longer after CPT-11 administration than after treatment with SN38 and suggested that clinicians should maintain plasma levels of SN38 to enhance the antitumor activity of CPT-11. One of the advantages of present invention provides clinicians with the ability to directly adjust the plasma levels of 11, 7-HECPT or 11,7-HMCPT derivatives to the point of therapeutic tolerance by controlling the dose and schedule of administration, which should lead to a superior ability to achieve better anti-tumor activity and reduce interpatient variability of the plasma levels of 11, 7-HECPT or 11, 7-HMCPT.
The different types of observations made in these studies suggest that direct administration of 11,7-HECPT or 11, 7-HMCPT by parenteral and oral administration could provide significant clinical benefit for patients with cancer which is amenable to treatment with this type of agent. However, in the past, similar hydrophobic camptothecin derivatives have been considered insufficiently water soluble for clinical use. The current invention overcomes this problem by providing lactone stable pharmaceutically acceptable multisolvent formulations of 11,7-HECPT for parenteral use and also oral 11,7-HECPT formulations.
This invention teaches novel 11-hydroxy modified camptothecins with 7-alkyl or 7-alkoxy modifications as new compositions of matter. These novel compounds which contain the 11-hydroxy or 11-alkoxy modifications of 7-alkyl or 7-alkoxy camptothecins represent new camptothecin derivatives, which are useful for the treatment of human cancers. Like many of the camptothecins, these molecules are generally poorly soluble in water and are more lipid soluble. Greater lipid solubility will facilitate diffusion of these molecules and permit more effective tissue penetration than the water soluble camptothecin species, such as CPT-11, E-ring opened carboxylate forms of camptothecin, or cationic species of camptothecins. To the inventor's knowledge, neither 11-hydroxy-7-ethyl camptothecin (11,7-HECPT), 11-hydroxy-7-methoxy camptothecin (11,7-HMCPT) or 11-hydroxy-7-alkoxy camptothecin have been directly administered to human subjects for the purpose of inhibiting the growth of cancer cells. This invention overcomes the above mentioned limitations and claims novel pharmaceutically acceptable formulations of 11,7-HECPT, and 11,7-HMCPT, methods of administration of 11,7-HECPT and 11,7-HMCPT, and antitumor compositions comprising solutions of 11,7-HECPT and solutions of HMCPT. Additionally, this invention claims novel dosages, schedules of administration, and routes of administration for both 11,7-HECPT and 11,7-HMCPT formulations to humans with various forms of cancer.